Urinary Tract Infections Question

(5) nursing research articles that discuss the use of the chosen nursing intervention REVIEW ARTICLE
Cranberry Juice and Urinary Tract Infection
R. Raz,1,2 B. Chazan,1 and M. Dan3
1
Infectious Diseases Unit, Haemek Medical Center, Afula, 2Rappaport School of Medicine, Technion, Haifa, and 3Infectious Diseases Unit,
Wolfson Medical Center, Holon, Israel
In women with recurrent urinary tract infections
(UTIs), long-term antimicrobial prophylaxis is indicated [1]. This method is effective but can cause adverse
reactions and can increase emergence of antimicrobial
resistance [2, 3]. Therefore, the need for alternative
therapies for UTI prophylaxis is evident. Cranberries
are one nonantibiotic alternative.
THE FRUIT
The scientific name for cranberry plant is Vaccinium
macrocarpon [4]. Cranberries, blueberries, and Concord
grapes are the only 3 fruits that are native to the United
States and Canada. Most commercial farms today are
located in northern United States, Massachusetts, and
New Jersey and the Canadian provinces of Quebec and
British Columbia [5]. Commercial harvests occur in
September and October.
Cranberries contain 180% water and 10% carbohydrates [6]. Among other constituents are flavonoids,
anthocyanins, catechin, triterpenoids, organic acids,
Received 4 November 2003; accepted 12 January 2004; electronically published
26 April 2004.
Reprints or correspondence: Dr. Raul Raz, Infectious Diseases Unit, Haemek
Medical Center, Afula, Israel (raz_r@clalit.org.il).
Clinical Infectious Diseases 2004; 38:1413–9
 2004 by the Infectious Diseases Society of America. All rights reserved.
1058-4838/2004/3810-0015$15.00
and a small amount of ascorbic acid. The major organic
acids are citric, malic, and quinic acids, with small
amounts of benzoic and glucuronic acids [7]. Anthocyanin pigments obtained from cranberry pulp are used
for coloring applications [8]. Cranberries can be processed into fresh fruit, concentrate, sauce products, and
juice drinks [5]. The single-strength juice is very acidic
(pH, !2.5) and unpalatable. In 1930, cranberry juice
cocktail, comprising a mixture of cranberry juice,
sweetener, water, and added vitamin C, was introduced.
The leading brand of cocktail contains 33% pure cranberry juice. Dried cranberry powder formulated in capsules or tablets is also available.
MECHANISTIC STUDIES:
URINE ACIDIFICATION
Native Americans were the first to use cranberries for
their medicinal properties [5]. Cranberries were used
for a variety of complaints, including blood disorders,
stomach ailments, liver problems, and fever. During the
1880s, German physicians observed that urinary excretion of hippuric acid increased after ingestion of
cranberries. In 1914, Blatherwick [9] published an article showing that cranberries are rich in benzoic acid,
which is then excreted in urine as hippuric acid. Therein
followed a long period during which the usefulness of
cranberry juice was thought to be based on the urinary
excretion of hippuric acid, which is a bacteriostatic
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Cranberries have long been the focus of interest for their beneficial effects in preventing urinary tract infections
(UTIs). Cranberries contain 2 compounds with antiadherence properties that prevent fimbriated Escherichia
coli from adhering to uroepithelial cells in the urinary tract. Approximately 1 dozen clinical trials have been
performed testing the effects of cranberries on the urinary tract. However, these trials suffer from a number
of limitations. Most importantly, the trials have used a wide variety of cranberry products, such as cranberry
juice concentrate, cranberry juice cocktail, and cranberry capsules, and they have used different dosing regimens. Further research is required to clarify unanswered questions regarding the role of cranberries in
protecting against UTI in general and in women with anatomical abnormalities in particular.
MECHANISTIC STUDIES: ANTIADHERENT
PROPERTIES
Adherence of uropathogens to uroepithelial cells is the initial
step in pathogenesis of UTI [20]. In 1984, Sobota [21] was first
to suggest that “reported benefits derived from the use of cranberry juice may be related to its ability to inhibit bacterial
adherence” (p. 1013). Sobota found that cranberry juice cocktail reduced adherence by 175% in 160% of 77 clinical isolates
of E. coli recovered from patients with UTI. Fifteen of 22 subjects showed significant antiadherence activity in their urine 1–
1414 • CID 2004:38 (15 May) • Raz et al.
3 h after drinking 15 oz (443.6 mL) of cranberry juice cocktail
[22].
Since Sobota’s initial report, several studies have confirmed
that the presumed efficacy of cranberry in preventing UTI is
related to its antiadherent properties. It is now known that E.
coli, the most common cause of UTI, have hairlike fimbria that
protrude from their surface. The fimbriae produce 2 adhesins
(mannose sensitive and mannose resistant) that attach to receptors on uroepithelial cells [23].
Zafriri et al. [24] identified 2 compounds in cranberries that
inhibit E. coli adhesins. One is fructose, which inhibits the
mannose-sensitive fimbrial adhesins; the other is a highmolecular-weight compound that inhibits the mannose-resistant adhesins of uropathogenic E. coli [25]. Although all fruit
juices contain fructose, only juices from Vaccinium berries (i.e.,
cranberries and blueberries) contain this second unique polymeric compound [26], which was later named “proanthocyanidin.” Interestingly, proanthocyanidin shows a very strong
inhibitory activity against mannose-resistant adhesins produced
by urinary isolates of E. coli [25] but shows only moderate
antiadherent activity against fecal E. coli isolates [27].
The antiadhesive property of cranberries probably helps to
prevent UTI in 2 ways: first, it directly prevents E. coli from
adhering to uroepithelial cells; and second, it selects for less
adherent bacterial strains in the stool. A recent study showed
that regular consumption of cranberry juice was also effective
in cases in patients with UTI caused by antibiotic-resistant
bacteria [28]. Urine samples obtained from healthy volunteers
who drank cranberry juice prevented uropathogenic E. coli isolates from adhering to isolated uroepithelial cells in bioassays.
The antiadherent effect started within 2 h and persisted for up
to 10 h after ingestion [29].
CLINICAL STUDIES
UTI prophylaxis. The first clinical study evaluating the effect
of cranberry on urinary tract was published in 1966. Papas et
al. [30] described the effect of cranberry juice in 60 patients
with bacteriuria who received 480 mL of juice daily for 3 weeks.
After therapy, 53% had a positive response and an additional
20% had a more modest benefit, but 6 weeks after stopping
treatment, bacteriuria reappeared in most of the subjects.
Since the study of Papas et al. [30], about a dozen clinical
trials evaluating various cranberry products have been performed. All these subsequent trials have studied the effect of
cranberry in preventing urinary tract symptoms. In some of
them, the primary parameter tested was UTI; in other studies,
bacteriuria was the primary end point. These trials have evaluated various patient populations, including sexually active
adult women, elderly or pediatric patients, and patients with
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agent and has the potential to acidify urine [10]. In 1923,
Blatherwick and Long [11] reported a reduction in urine pH
level (6.4–4.5) with a concomitant increase in excretion of hippuric acid (0.77–4.74 g) after subjects ate 350 g of cooked
cranberries [12]. In 1933, Fellers et al. [13] published results
for 6 men who ingested 100–300 g of cranberries daily. They,
too, showed an increase in acidity and excretion of organic
acids (including hippuric acid) in urine. However, these authors
concluded that an ordinary serving of 22–54 g of cranberries
produced only a very slight increase in urine acidity [14].
Kinney and Blount [15] found that certain amounts of cranberry juice (450–720 mL daily) lowered urinary pH. Similarly,
Jackson and Hicks [16] showed that 710 mL of cranberry juice
(25% pure juice) lowered the urine pH of 21 elderly men. In
contrast, Nahata et al. [17] reported that the addition of vitamin
C to cranberry cocktail did not change urine pH. Notably, an
early study by Bodel et al. [18] in 1959 found that, in 5 healthy
subjects, 1200–1400 mL of cranberry juice cocktail slightly acidified urine and increased the amount of hippuric acid excreted
in urine to 3–4 g. However, none of the urine samples from
these patients was bacteriostatic against Escherichia coli. In addition, Bodel et al. [18] demonstrated that hippuric acid was
bacteriostatic at a minimum concentration of 0.02 mmol/L at
pH 5.0, and the antibiotic activity of hippuric acid decreased
∼5-fold as the pH increased to 5.6. Bodel et al. [18] concluded
that cranberry juice could not exert a bacteriostatic effect because it was not a rich enough source of hippuric acid, coupled
with the fact that it did not lower urine pH sufficiently.
The validity of the conclusions of Bodel et al. [18] was subsequently confirmed by others. Today, it is known that the low
amount of benzoic acid present in the fruit (!0.1% of weight),
coupled with maximum tolerated amounts of cranberry juice
(∼4 L/d), rarely results in enough hippuric acid excretion necessary to achieve bacteriostatic urinary concentrations [19]. Ingestion of large amounts of cranberry juice is required to
slightly reduce pH of urine and modestly increase hippuric acid
excretion, changes that do not confer significant antibacterial
activity to urine. If cranberry juice is a protective agent for the
urinary tract, then another mechanism must be involved.
drinking of cranberry juice was associated with decreased risk
of UTI. Although this study was retrospective and examined
first-time UTI and not recurrent UTI, it adds to the notion
that young, sexually active women constitute a population that
may benefit from cranberry products.
Two studies have evaluated the use of cranberries in elderly
women, but unlike the above 3 prospective studies, these trials
chose bacteriuria as their primary parameter. Avorn et al. [38]
conducted a large randomized, double-blind study in which
153 asymptomatic elderly women received 300 mL per day of
cranberry juice cocktail or placebo. Urine samples were obtained at baseline and at 1-month intervals for 6 months, and
tested for bacteriuria and pyuria. At baseline, bacteriuria and
pyuria were present in ∼20% of samples in both the cranberry
group and the placebo group. At the 1-month mark, there was
no difference in the percentage of urine samples with bacteriuria and pyuria in the 2 groups (∼25%). However, from
the 2-month mark on, there was a statistically significant difference between groups favoring the cranberry group. At the
end of the 6-month study, bacteriuria and pyuria were present
in 28% of urine samples from the cranberry group. The chances
of having bacteriuria with pyuria were 42% less in the cranberry
group than in the control group. These authors concluded that
ingesting cranberry beverages reduced the frequency of bacteriuria with pyuria in older women, although they noted that,
in elderly women, asymptomatic bacteriuria does not usually
require treatment. Nevertheless, in their study, there were 16
instances of antibiotic use for UTI in the control group versus
8 in the cranberry group.
Haverkorn and Mandigers [39] also evaluated the use of
cranberry by elderly patients, but they used a nonblinded crossover design. Men and women in a nursing department of a
general hospital were provided 15 mL cranberry juice (type not
detailed) mixed with water twice daily or the same amount of
water daily. After 4 weeks, the regimens were reversed. Only
17 patients stayed in the department long enough to complete
both 4-week periods. Bacteriuria was observed in 3 patients
during the entire time course and in neither period in 7 patients.
In the additional 7 patients, there were fewer instances of bacteriuria during the cranberry period than during the control
period, supporting a moderately preventive role for cranberry
juice.
Two additional but not randomized trials involving elderly
patients were conducted. In a Danish trial [33], the incidence
of UTI was compared in 2 geriatric departments. Patients were
offered cranberry juice in one department and the usual mixed
berry juice in the other. The results showed that cranberry juice
did not influence incidence of UTI. In another study, 538 nursing home residents were provided either 220 mL of cranberry
juice or 6 capsules containing cranberry extract daily [36].
Compared with historical controls, the incidence of UTI was
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different medical conditions. Table 1 summarizes the relevant
prospective clinical studies conducted with cranberry products.
Kontiokari et al. [32], Stothers [31], and Walker et al. [35]
published randomized studies that examined adult women. In
the study by Kontiokari et al. [32], which was an open, randomized, controlled trial, 150 women were divided into 3
groups: one group drank 50 mL a day of cranberry-lingonberry
juice concentrate containing 7.5 g cranberry concentrate and
1.7 g lingonberry concentrate (lingonberry is another fruit of
the Vaccinium genus); another group drank 100 mL of a lactobacillus drink; and a third group received no intervention.
After 6 months’ treatment, 16% of the cranberry group, 39%
of the lactobacillus group, and 36% of the control group had
experienced ⭓1 recurrence of UTI. This translates to a 20%
reduction in absolute risk for the cranberry group. Interestingly,
even though the cranberry group stopped their treatment after
6 months (because the manufacturer stopped producing the
juice), the percentage of women who experienced recurrence
at 12 months was still significantly lower in the cranberry group,
implying a residual effect supporting the hypothesis that cranberry selects for less adherent bacterial strains.
Stothers [31] performed a randomized, placebo-controlled,
double-blind study. A total of 150 women with previous UTI
were divided into 3 groups: persons who received placebo juice
and placebo tablets, persons who received cranberry juice and
placebo tablets, and persons who received placebo juice and
cranberry tablets. Juice was 250 mL of pure unsweetened cranberry juice taken 3 times daily, and tablets contained 1:30 parts
concentrated juice taken twice daily. After 1 year, results showed
that 32% of placebo recipients had experienced ⭓1 UTI during
the year, compared with 20% in the cranberry juice group and
18% in the cranberry tablet group. The absolute risk reduction
for cranberry products was 12%–14%, similar to the findings
of Kontiokari et al. [32].
A smaller study by Walker et al. [35] adds further support
to the above findings. In this study, which followed a doubleblind crossover design, 19 women with recurrent UTI were
provided either cranberry capsule (with 400 mg cranberry solids) or a placebo capsule for 3 months. Patients then switched
to an alternative therapy for the next 3 months. Although only
10 patients finished the entire course of treatment, results favored the use of cranberry. Of 21 episodes of UTI, 6 occurred
in the cranberry group and 15 occurred in the placebo group.
Overall, these 3 studies show that use of cranberry is effective,
at least statistically, for prophylaxis of UTI in adult women with
recurrent UTI. It should be noted, however, that none of these
3 studies used the popular commercial brand of cranberry juice
cocktail.
Of interest too is an epidemiological study [40] that evaluated
the relationship between health/sexual behavior and first-time
UTI in sexually active women. The study found that regular
Year of
study
2002
2001
2001
1999
1997
1997
1995
1994
1994
1966
Stothers et al. [31]
Kontiokari et al. [32]
Kirchhoff et al. [33]
Schlager [34]
Walker et al. [35]
Dignam et al. [36]
Foda [37]
Avorn [38]
Haverkorn and Mandigers [39]
Papas et al. [30]
Population
2 geriatric units
150 women with previous UTI
150 women with previous UTI
Nonrandomized
Quasi randomized cross-over
220 mL of cranberry juice or 6 capsules
with cranberry extract per day
Cranberry capsule with 400 mg
of cranberry solids vs. placebo, each
for 3 months
60 patients (73% women)
with bacteriuria
38 elderly men and women
(17 finished the study)
Bacteriuria and pyuria were significantly reduced:
28% of samples from placebo recipients vs.
15% of samples from cranberry patients
No benefit in preventing UTI or bacteriuria
Compared with historical controls, incidence
of UTI significantly reduced, from 27 cases
per month to 20 cases per month
Cranberry effective in preventing UTI: of 21
UTIs, 6 UTIs were in the cranberry group
and 15 were in the placebo group
480 mL of cranberry juice for 21 days
Positive response in 53% of cases and modest
response in 20%
15 mL of cranberry juice mixed with water 7 of 17 patients had reduction of bacteriuria
b.i.d. vs. water, each for 4 weeks
during cranberry period
300 mL of cranberry juice cocktail vs.
placebo for 6 months
40 children with neurogenic bladder Cranberry cocktail, 15 mL/kg/d, vs. water,
(21 finished)
each for 6 months
538 nursing home residents
Quasi randomized placebo-controlled, 153 elderly women
double-blind
Randomized, single-blind, cross-over
Nonrandomized, historical controls
Randomized, double blind, cross-over 19 women with recurrent UTI
(10 finished the study)
No benefit in preventing UTI or bacteriuria
No effect on UTI
A significant reduction in UTI: 16% for cranberry
vs. 39% for lactobacillus and 36% for no
intervention
50 mL cranberry-lingonberry concentrate
vs. 100 mL lactobacillus drink vs. no
intervention for 6 months
Cranberry juice vs. usual mixed berry
juice; mean stay, 4 weeks
Outcome
A significant reduction in UTI: 18% for cranberry
tablets vs. 20% for cranberry juice vs. 32%
for placebo
Intervention
Placebo juice/tablets vs. placebo juice/
cranberry tablets vs. cranberry juice/
placebo tablets; tablets were given
b.i.d., and juice was given as 250 mL
pure unsweetened product, t.i.d.; 1 year
trial
Randomized, double-blind, cross-over 15 children with neurogenic bladder 300 mL cranberry concentrate vs.
placebo, each for 3 months
Nonrandomized, controlled
Open, randomized
Randomized, placebo-controlled,
double-blind
Method
Summary of prospective studies evaluating prophylaxis of urinary tract infection (UTI) on bacteriuria.
Study
Table 1.
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trial. Another nonrandomized study [47] found decreased leukocyte counts in urine samples obtained from handicapped
children (most with indwelling catheters) who drank cranberry
juice. This, too, was not a randomized trial.
UTI treatment. The Cochrane reviewers concluded that
randomized studies assessing effectiveness of cranberry juice
for treatment of UTI have not yet been conducted. Therefore,
at present, there is no evidence to suggest that cranberry juice
or other cranberry products are effective for treatment of UTI.
The safety of cranberries is considered to be excellent. Some
patients may experience a slight laxative effect, depending on
the amount ingested [9, 15, 16]. Nevertheless, at least one author has warned that ingesting a large amount of cranberries
over a long duration may increase risk of some types of urinary
stones in high-risk patients because of the increased urinary
excretion of oxalate and slight urinary acidification [48].
CONCLUSION AND DIRECTIONS
FOR THE FUTURE
Results of clinical studies suggest a possible clinical benefit of
cranberry juice in preventing UTI in some populations. The
strongest evidence available is for sexually active adult women
with previous UTI. In this population, cranberry appears to be
effective in the prophylaxis of recurrent UTI, although standard
juice cocktail was not specifically tested. In elderly patients,
cranberry consumption reduces the incidence of bacteriuria,
although this is often not treated with antibiotics. In contrast,
none of the randomized clinical studies that evaluated patients
at high risk of UTI—for example, those with neurogenic bladder—found cranberries to have a beneficial effect.
In the population that benefits most from the prophylactic
effect of cranberry intake (sexually active women with recurrent
UTI), trial results repeatedly show an ∼50% reduction in disease
morbidity. From a clinical point of view, this is quite a modest
benefit, considering the accompanying burden of long-term
daily intake of the compound. Not less significant is the inconvenience associated with the amount of juice required to
assure continuous availability and the need to carry a daily
supply if twice- or thrice-daily dosing is needed to work, business, or vacation travel. If one considers the understandably
high rate of dropouts, the 50% efficacy rate may drop to a
remarkably lower effectiveness.
Furthermore, results of the reviewed studies should not be
viewed as conclusive because many of the trials suffer from
various limitations, including lack of randomization, no or improper blinding, small number of subjects, short trial duration,
large number of dropouts, and no reported intent-to-treat
analysis [43]. Perhaps the single most consistent limitation of
these trials is the lack of uniformity regarding the intervention,
including the particular cranberry product evaluated (juice,
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significantly reduced, from 27 cases a month to 20 cases a
month.
Two studies have evaluated the potential of cranberry in
pediatric patients with medical conditions predisposing them
to UTI. These trials did not show any benefit of cranberry for
prevention of UTI or bacteriuria. In the crossover, placebocontrolled, double-blind study of Schlager et al. [34], 300 mL
of cranberry concentrate provided for 3 months did not have
any benefit when provided to 15 children with neurogenic bladder receiving intermittent catheterization. Frequency of bacteriuria was 75% during both placebo and cranberry periods,
and the number of UTIs was not significantly different either.
In a similar patient population, Foda et al. [37] administered
water or cranberry cocktail (15 mL/kg/d) to 40 children for 6
months, and the reverse for an additional 6 months. Only 21
patients finished the study. Cranberry had no effect on the
frequency of UTI or bacteriuria. Two additional studies were
performed in adult patients, but neither study evaluated clinical
outcomes. In 8 adult patients with multiple sclerosis randomized to receive 20 days’ therapy, cranberry increased acidity of
urine, and in 15 patients with spinal cord injury, cranberry
reduced bacterial biofilm load in the bladder [41, 42].
Jepson et al. [43] reviewed in the Cochrane Library all randomized or quasi-randomized controlled studies for the prevention of UTI with cranberry juice. Until 2000, only 5 trials
met all the criteria adopted for evaluation. (These 5 trials are
discussed above.) The conclusion of the review was that because
of the small number and poor quality of trials, there is insufficient evidence to show the effectiveness of cranberry juice for
prevention of UTI. However, the Cochrane reviewer did not
include the latest 2 studies by Stothers [31] and Kontiokari et
al. [32]. Both studies were randomized and large, and they
found that women with previous UTIs who took cranberry
products as prophylaxis experienced fewer recurrent UTI. On
the basis of these 2 trials, a recent evidence-based answer published in the Journal of Family Practice [44] suggested that a
trial of cranberry juice (3 glasses daily) was reasonable for
women with recurrent UTI who are being considered for antibiotic prophylaxis. The author of the “answer” also noted that
“no national practice guidelines have recommended cranberry
juice as a preventive strategy for recurrent UTI” [44, p. 155].
However, educational brochures published by the National Kidney Foundation contain statements supporting the possible use
of cranberry juice in helping to prevent the development of
UTI [45].
Evidence regarding the role of cranberries for treating, rather
than preventing, UTI is almost nonexistent. The same Cochrane
reviewer who evaluated UTI prevention also systematically reviewed the literature for trials that evaluated use of cranberries
for treating UTI [46]. As mentioned above, Papas et al. [30]
studied patients with bacteriuria, but this was not a randomized
Acknowledgments
We thank Frances Nachmani, Hana Edelstein, and Miriam
Ben-Yashar for their assistance in data collection and processing.
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sweetened cocktail, or capsules/tablets), concentration, dosing
regimen, and duration of the intervention, which greatly differed from study to study [43, 49]. Further properly designed
trials addressing these issues are needed [38].
Future trials should also assess patient acceptability of treatment. Some studies have indicated that the taste and caloric
load of cranberry juice cocktail is unacceptable to many patients, especially over the long term [15, 40, 44]. Capsules of
cranberry concentrate could be a better-tolerated alternative
[31]. Cost is another issue that affects patient uptake of treatment, because cranberry products are not currently covered by
health insurance [44, 49]. In the single trial that evaluated the
issue of cost, Stothers [31] found that cranberry tablets are
more cost-effective than organic cranberry juice.
Therefore, the potential of cranberry products to act as a
nonantibiotic alternative for preventing UTI, thereby reducing
the total amount of antibiotics prescribed for UTI, could have
great public health significance. In November 2002, the National Center for Complementary and Alternative Medicine, a
branch of the US National Institutes of Health, announced an
initiative to fund research on the role of cranberry in promoting
urinary tract health [50]. As antimicrobial resistance continues
to climb, the time is ripe to recognize the importance of further
cranberry research.
38.
39.
40.
41.
42.
JF. Efficacy of cranberry in prevention of urinary tract infection in a
susceptible pediatric population. Can J Urol 1995; 2:98–102.
Avorn J, Monane M, Gurwitz JH, Glynn RJ, Choodnovskiy I, Lipsitz
LA. Reduction of bacteriuria and pyuria after ingestion of cranberry
juice. JAMA 1994; 271:751–4.
Haverkorn MJ, Mandigers J. Reduction of bacteriuria and pyuria using
cranberry juice [letter]. JAMA 1994; 272:590.
Foxman B, Geiger AM, Palin K, Gillespie B, Koopman JS. First-time
urinary tract infection and sexual behavior. Epidemiology 1995; 6:
162–8.
Schultz A. Effect of cranberry juice and ascorbic acid in acidifying the
urine in multiple sclerosis subjects. J Community Health Nurs 1984;
1:159–69.
Reid G, Hsiehl J, Potter P, et al. Cranberry juice consumption may
reduce biofilms on uroepithelial cells: pilot study in spinal cord injured
patients. Spinal Cord 2001; 39:26–30.
43. Jepson RG, Milhaljevic L, Craig J. Cranberries for preventing urinary
tract infections. Cochrane Database Syst Rev 2001:CD001321.
44. Kiel R, Nashelsky J. Does cranberry juice prevent or treat urinary tract
infection? J Fam Pract 2003; 52:154–55.
45. National Kidney Foundation. Available at: http://www.kidney.org.
46. Jepson RG, Milhaljevic L, Craig J. Cranberries for treating urinary tract
infections. Cochrane Database Syst Rev 2000:CD001322.
47. Rogers J. Pass the cranberry juice. Nursing Times 1991; 87:36–7.
48. Terris MK, Issa MM, Tacker JR. Dietary supplementation with cranberry concentrate tablets may increase the risk of nephrolithiasis. Urology 2001; 57:26–9.
49. Harkins KJ. What’s the use of cranberry juice? Age Ageing 2000; 29:
9–12.
50. US National Institutes of Health. National Center for Complementary
and Alternative Medicine. Available at: http://nccam.nih.gov/research/
concepts/consider/cranberry.htm. Accessed on 25 July 2003.
Downloaded from https://academic.oup.com/cid/article/38/10/1413/345228 by guest on 24 April 2022
Cranberry and Urinary Tract Infection • CID 2004:38 (15 May) • 1419
MISCELLANEOUS
Prevention of Urinary Tract Infection with Oximacro®, A Cranberry Extract with a High Content of
A-Type Proanthocyanidins: A Pre-Clinical Double-Blind Controlled Study
Andrea Occhipinti,1 Antonio Germano,2 Massimo E. Maffei1*
Purpose: Urinary tract infections (UTIs) are widespread and affect a large portion of the human population. Cranberry juices and extracts have been used for UTI prevention due to their content of bioactive proanthocyanidins
(PACs), particularly of the A type (PAC-A). Controversial clinical results obtained with cranberry are often due
to a lack of precise determination and authentication of the PAC-A content. This study used Oximacro® (Biosfered S.r.l., Turin, Italy), a cranberry extract with a high content of PAC-A, to prevent UTIs in female and male
volunteers.
Materials and Methods: The Oximacro® PACs content was assayed using the Brunswick Laboratories 4-dimethylaminocinnamaldehyde (BL-DMAC) method, and the dimer and trimer PACs-A and PACs-B percentages
were determined via high-performance liquid chromatography/electrospray ionization tandem mass spectrometry
(HPLC/ESI-MS/MS). A balanced group of female (ranging from 19 to over 51 years) and male volunteers (over
51 years) was divided into two groups. The experimental group received 1 capsule containing Oximacro® (36 mg
PACs-A) twice per day (morning and evening) for 7 days, and the placebo group was given the same number of
capsules with no PACs.
Results: Analysis of Oximacro® revealed a high total PAC content (372.34 mg/g ± 2.3) and a high percentage of
PAC-A dimers and trimers (86.72% ± 1.65). After 7 days of Oximacro® administration, a significant difference
was found between the placebo and Oximacro® groups for both females (Mann-Whitney U-test = 875; P < .001; n = 60) and males (Mann-Whitney U-test = 24; P = .016; n = 10). When the female and male age ranges were analysed separately, the female age range 31-35 showed only slightly significant differences between the placebo and Oximacro® groups (Mann-Whitney U-test = 20.5; P = .095; n = 10), whereas all other female age ranges showed highly significant differences between the placebo and Oximacro® groups (Mann-Whitney U-test = 25; P = .008; n = 10). Furthermore, colony forming unit/mL counts from the urine cultures showed a significant difference (P < .001) between the experimental and the placebo groups (SD difference = 51688; df = 34, t = -10.27; Dunn-Sidak Adjusted P < .001, Bonferroni Adjusted P < .001). Conclusion: Careful determination of the total PAC content using the BL-DMAC method and the authentication of PACs-A with mass spectrometry in cranberry extracts are necessary to prepare effective doses for UTI prevention. A dose of 112 mg Oximacro® containing 36 mg PACs-A was found to be effective in preventing UTIs when used twice per day for 7 days. Keywords: urinary tract infections; prevention & control; plant extracts; pharmacology; humans; urinalysis; therapeutic use; Vaccinium macrocarpon. INTRODUCTION U rinary tract infections (UTIs) are widespread and affect a large portion of the human population. Approximately 13 million women in the United States and approximately 150 million people worldwide develop UTIs each year, with societal costs of approximately 3.5 billion USD per year in the USA alone.(1) An estimated 40% of women develop at least one UTI during their lifetimes.(2) UTIs refer to the presence of a certain threshold number of bacteria in the urine (usually > 105/mL) and consist of cystitis (or lower UTIs,
with bacteria in the bladder), urethral syndrome and
pyelonephritis (or upper UTIs, with infection of the
kidneys).(3) Bacterial cystitis (also called acute cystitis)
can occur in women and men, and the signs and symptoms include dysuria (pain on passing urine), frequency, cloudy urine, and occasionally haematuria (blood in
the urine); bacterial cystitis is also often associated with
pyuria (urine white cell count > 104/mL). Some people
also develop recurrent UTIs with an average of two to
three episodes/year.(4)
The berries of cranberry (Vaccinium macrocarpon Ai-
1
Department of Life Sciences and Systems Biology, University of Turin, Via Quarello 15/A, Turin, Italy.
Farmacia Antoniana, Viale Cesare Balbo, 3, 10040 San Gillio (TO), Italy.
*Correspondence: Department of Life Sciences and Systems Biology, University of Turin, Via Quarello 15/A, 10135 Turin,
Italy.
Tel: +39 011 6705967. Fax: +39 011 2365967. E-mail: massimo.maffei@unito.it.
Received September 2015 & Accepted February 2016
2
Vol 13 No 02 March-April 2016 2640
2590
Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al.
Figure 1. Spectral analysis of the BL-DMAC reaction of PAC-A (black line) and PAC-B (red line). When tested at the same concentration (20 µg mL-1),
PAC-B showed a higher absorbance (at 640 nm) than PAC-A.
Abbreviations: PAC, proanthocyanidins; BL-DAMC, Brunswick Laboratories 4-dimethylaminocinnamaldehyde.
ton) have been used for hundreds of years as a remedy
for diseases of the urinary tract and have attracted attention due to their potential health benefits.(5,6) The beneficial mechanism of cranberry was historically thought to
be due to the fruit’s acids causing a bacteriostatic effect
in the urine. However, recently, a group of proanthocyanidins (PACs) with A-type linkages (PAC-A) was
isolated from cranberries and shown to exhibit bacterial
antiadhesion activity against both antibiotic-susceptible and -resistant strains of uropathogenic P-fimbriated
Escherichia coli (E. coli) bacteria, including multidrug-resistant E. coli.(7-10) Central in the efficacy of cranberry extract/juice is the determination of the optimum
dose of PAC-A, which is an essential requirement in
establishing botanical supplements as viable supports
to conventional therapies.(11) Recent studies have revealed that cranberry extract regimens containing 72
mg PACs produce significant bacterial antiadhesion activity in human urine.(2,10) Clearly, the dose of bioavailable PAC-A is central to the issue of cranberry efficacy.
Currently, four methods are used to evaluate the content of cranberry PACs; two methods are based on the
depolymerisation of PACs (e.g., the hydrochloric acid
butanol method known as Bates-Smith and the European Pharmacopoeia method), and two are colorimetric
methods (a ultraviolet-visible [UV-VIS] spectrophotometric method based on Prussian-blue or Folin-Ciocalteu reagents and the Brunswick Laboratories 4-dimethylaminocinnamaldehyde [BL-DMAC] method). The
BL-DMAC colorimetric method (an aldehyde condensation of 4-dimethylaminocinnamaldehyde) appears to
be more accurate than the other methods and has been
successfully used to quantify cranberry PACs.(12) In parMiscellaneous 2641
ticular, the BL-DMAC method is less likely to be subject to interference from cranberry components, such as
anthocyanins, because the reaction is read at 640 nm.
However, the BL-DMAC method, although specific for
PAC quantification, is not able to distinguish between
A- and B-type PACs;(13) therefore, analytical methods, such as high-performance liquid chromatography
(HPLC) coupled to mass spectrometry or fluorescence
detectors, are necessary for PACs-A authentication.(14)
Oximacro® is a cranberry extract with the highest content of PACs (according to the BL-DMAC method) and
the highest percentages of PAC-A dimers and trimers
(based on Liquid chromatography [LC]/mass spectrometry [MS] identification) available on the market. Here,
we report on the chemical analysis of the PAC content
of Oximacro® and its action in preventing UTIs based
on a pre-clinical double-blind controlled study on male
and female volunteers.
MATERIALS AND METHODS
Reagents
Oximacro®, a cranberry (Vaccinium macrocarpon
Aiton) extract, was provided by Biosfered S.r.l. (Turin, Italy) and produced from cranberries as a reddish
powder with a total PAC content > 360 mg/g (Lot #
CR0104-PD01). The CoA of the product is available at
the company web site (http://www.biosfered.com). Extrasynthese (Lyon, France) provided pure standards of
PAC-A and PAC-B. The pure chemicals were dissolved
in 96% v/v ethanol (Sigma-Aldrich, Carlsbad, USA) at
a final concentration of 100 µg/mL. Aliquots of stock
solutions were stored in 1.5-mL HPLC vials at -80°C
Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al.
Figure 2. Time-course of the BL-DMAC reaction of 20 µg mL-1 PAC-A and PAC-B. When used at the same concentration, PAC-B (dotted line) reacts
more rapidly and with a higher absorbance with respect to PAC-A (solid line). Metric bars represent standard deviation.
Abbreviations: PAC, proanthocyanidins; BL-DAMC, Brunswick Laboratories 4-dimethylaminocinnamaldehyde.
until use. The chemical purity and integrity of standard
compounds was assessed (see below) prior to use.
Determination of the Total PAC Content with the
BL-DMAC Method
The BL-DMAC assay was performed according to the
method of Prior and colleagues(12) with minor modifications. Extraction buffer was composed of acidified 75%
v/v acetone (VWR International, Milan) with 0.5% v/v
acetic acid (Sigma-Aldrich, Carlsbad, USA). Acidified
ethanol was composed of 72% v/v ethanol and hydrochloric acid (Sigma-Aldrich, Carlsbad, USA) at a final
concentration of 1.52 M. DMAC solution was composed of 0.1% w/v 4-(dimethylamino)-cinnamaldehyde
(DMAC) (Sigma-Aldrich, USA) in acidified ethanol;
this solution was freshly prepared prior to the assay.
Briefly, Oximacro® (20-30 mg) was dissolved in 5 mL
of extraction buffer. The powder was extracted in an
ultrasonic bath at room temperature for 20 min and then
shaken with an orbital shaker for 1 h. Samples were
centrifuged at 5,000 g for 10 min and then diluted in the
extraction buffer prior to spectrophotometric assay. The
colorimetric reaction was performed by mixing 0.84
mL DMAC solution and 0.28 mL of a diluted sample in
a 1.5-mL plastic cuvette. The total PACs were quantified via an external calibration curve made with a pure
PAC-A standard. The reaction kinetics of both PAC
standards (PAC-A and PAC-B) were determined using
a time-course BL-DMAC assay. A concentration of 20
µg/mL was tested for both standards. The reaction was
incubated in the dark from 1 to 25 min to assess the dynamics of the DMAC reaction, and the absorbance was
read at 640 nm (Cary60, Agilent-Technologies, Califor-
nia, USA) against a blank composed of acidified ethanol
and DMAC solution. The quantification was performed
in triplicate within the linear range of calibration curves
(5-30 µg/mL). Oximacro® was then assayed exactly
at 20 min, which corresponds to the maximum absorbance value for PAC-A. To test the reactivity of PAC-A
and PAC-B to DMAC, 20 µg/mL solutions of PAC-A
and PAC-B were tested with increasing percentages of
PAC-B (0, 25, 50, 75 and 100%). The final concentration of the tested mixtures was always 20 µg/mL. The
absorption spectra were recorded between 350 and 800
nm, exactly 20 min after the beginning of the DMAC
colorimetric assay.
Authentication of the PAC-A Content in Oximacro®
PAC-A and PAC-B authentication of Oximacro® were
obtained via liquid chromatography (1200 HPLC, Agilent Technologies, California, USA) equipped with a
reverse phase (RP) C18 Kinetex (2.6 μm, 100 × 3.0
mm, Phenomenex, California, USA) column. The binary solvent system was A) MilliQ H2O (Millipore,
Billerica, Massachusetts, USA) with 0.1% v/v of formic acid and B) acetonitrile (VWR International, USA)
with 0.1% v/v of formic acid. Chromatographic separation was carried out at constant flow rate (200 µL/min)
using the following conditions: linear gradient from 5%
to 30% of B in 10 min and isocratic elution for 5 min
and 20 min at 50% of B and 24 min at 90% of B at
24 min. The initial mobile phase was re-established for
10 min prior to the next injection. Tandem mass spectrometry analyses were performed with a 6330 Series
Ion Trap LC-MS System (Agilent Technologies, California, USA) equipped with an electrospray ionization
Vol 13 No 02 March-April 2016 2642
Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al.
Figure 3. BL-DMAC spectral analysis of a 20 µg mL-1 solution of PAC-A and PAC-B used at increasing PAC-B concentrations. Increasing PAC-B
causes an absorbance increase at 640 nm.
Abbreviations: PAC, proanthocyanidins; BL-DAMC, Brunswick Laboratories 4-dimethylaminocinnamaldehyde; Abs, absorbency.
source (ESI) operating in negative mode. The identification of PAC-A (dimers and trimers) was performed
via multiple reaction monitoring (MRM) by monitoring
the following parental ions [M-H]-: 575, 577, 861, 863
and 865 m/z.
Study Population and Inclusion and Exclusion
Criteria
To assess the effect of Oximacro®, we recruited participants from a population of volunteers (10 male and 60
female) involved in studies performed by the Farmacia
Antoniana (San Gillio, Italy) under the supervision of
medical doctors. Informed consent was obtained. The
inclusion criteria included any woman or man at least
18 years of age to over 51 years of age with at least 2
culture-documented symptomatic UTIs in the calendar
year prior to recruitment. The choice of volunteers was
completely balanced, and volunteers with known anatomical abnormalities (posterior urethral valves, neurogenic bladder, or any urinary obstruction) were excluded from this study. Urinary infection was defined as a
positive culture of a midstream sample with a uropath-
ogenic bacterium at 105 colony forming unit (CFU)/mL
in symptomatic volunteers with no more than two species of organisms present. We accepted lower counts
(104 CFU/mL) if the volunteer had typical symptoms
of UTI and positive white blood cells and/or nitrites on
urine analyses. Specific symptoms and signs included pain before, during, or after micturition; increased
frequency of micturition; pain in abdomen; haematuria; foul smell; and signs of common sickness (fever
> 37.9°C or 1.5°C above baseline, temperature, chills,
nausea, and vomiting). Nonspecific symptoms were
considered anorexia, fatigue and reduced mobility, and
signs of delirium (e.g., confusion and deterioration in
mental or functional status). Asymptomatic bacteriuria
was not considered an end point. After explaining the
study and obtaining consent, patients were assigned to
the placebo group or experimental randomized groups.
The randomization was concealed.
Oximacro® Administration and Dosage
Capsules contained 500 mg of the product [112 mg
Oximacro® (equivalent to 36% PAC-A), 383 mg mi-
Table 1. Comparative analysis of total PACs content based on a PAC-A calibration curve with an increasing percentage of PAC-B (± standard error; n
= 3). In the same column, different letters indicate significant (P < .05) differences. Percentage of a 20 µg mL-1 PAC-A Solution Percentage of a 20 µg mL-1 PAC-B Solution Total PACs, expressed as µg mL1 (± standard deviation) 100 0 20.49a (± 0.65) 75 25 22.65b (± 1.48) 50 50 25.49c (± 2.55) 25 75 28.18d (± 2.48) 0 100 31.29e (± 2.19) Abbreviations: PAC, proanthocyanidins. Miscellaneous 2643 Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al. Table 2. Volunteers baseline characteristics. Variables Experimental Group, Oximacro® Administration, n = 35 Placebo Group, n = 35 Demographics Females, no (%) 30 (85.7) 30 (85.7) Males, no (%) 5 (14.3) 5 (14.3) Median age (range) 38 (19-61) 38 (19-63) Age range 19-24 5 (F) 5 (F) 25-30 5 (F) 5 (F) 31-35 5 (F) 5 (F) 36-40 5 (F) 5 (F) 41-50 5 (F) 5 (F) > 51
5 (F), 5 (M)
5 (F), 5 (M)
Baseline characteristics
Acute UTI, no (%) 35 (100) 35 (100)
Bladder and bowel dysfunction, no (%)
35 (100)
35 (100)
Average UTIs in years prior to treatment
2.5
2.6
Number of capsules (days)
2 (7)
2 (7)
Volunteers not completing the study, no (%)
6 (17)
15 (43)
Females, no (%) 6 (17) 13 (43)
Males, no (%) 1 (20) 3 (60)
Abbreviations: M, male; F, female; UTI, urinary tract infections.
crocrystalline cellulose and 5 mg magnesium stearate].
The placebo was indistinguishable in colour, taste, and
appearance, consisting of all elements above without
Oximacro® and coloured with azorubine. The experimental group (5 males and 30 females) received 1 capsule containing 36% PAC-A twice per day (morning
and evening) for 7 days, and the placebo group (5 males
and 30 females) was given the same number of capsules
with no PACs. A score (from 0, representing no effect,
to 10, representing a maximum effect of Oximacro® in
preventing UTI) was recorded for all volunteers. To obtain linearity, the logarithm of the scores (Ln scores)
was used. The dose was calculated based on previous
clinical trials.(10) The administration was performed for
7 days; during this time, the volunteers were followed
with alternating visits and telephone calls every 2 days.
At the end of the treatment period, a urine sample was
sent for urine analysis and urine culture. To avoid conTable 3. Contingency table.
Variables Placebo Oximacro®
Not recovered 35 7
Recovered 0 28
Fisher’s Exact Test: P < .001 tamination, the volunteers were asked to not use antibiotics or any other cranberry products for the duration of the study, with the exception of the placebo group, in which volunteers were asked to immediately report on symptoms. In the latter case, they were asked to use the antibiotic prescribed by the medical doctor and to interrupt the placebo administration. The attending urologists, outcome assessor and statistician were all blinded to the group allocations. Statistical Analysis We performed Fisher’s exact tests on the tabulated frequencies to assess the effect of the treatments. Kolmogorov–Smirnov tests were used to assess the distribution type for the continuous variable, i.e., the average value of the score. The data were log-transformed. Accordingly, a non-parametric analysis of variance was used to assess the differences in the Oximacro® and placebo groups according to the sex and age categories. The median, quartile, maximum and minimum score values are represented in boxplots; outliers are represented by asterisks. A binary logistic regression was performed to test the independent effects of age and sex on Oximacro® outcomes. All statistical analyses were performed using Statistical Package for the Social Science (SPSS Inc, Chicago, Illinois, USA) version 22.0. The intenVol 13 No 02 March-April 2016 2644 Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al. Figure 4. HPLC-ESI-MS/MS analysis of Oximacro®. A) HPLC-ESI-MS/ MS chromatogram of dimers and trimers of PAC-A and PAC-B. The mass spectrometry analysis performed in MRM mode shows the presence of PAC-A dimers (black line) and trimers (blue line) and PAC-B dimers (red line). B) MS2 spectra in negative mode of a typical PAC-B dimer ([M-H]1 575 m/z). C) MS2 spectra in negative mode of a typical PAC-A dimer ([M-H]-1 577 m/z). Abbreviations: PAC, proanthocyanidins; HPLC-ESI-MS/MS, high-performance liquid chromatography/electrospray ionization tandem mass spectrometry; MRM, Multiple Reaction Monitoring. showed that the purity was identical to that declared by the supplier. To determine whether a mixture of PAC-A and PAC-B with increasing percentages of the two PACs could result in differing PAC quantifications, we assayed the BL-DMAC of the mixtures and found that the presence of PAC-B increased the absorbance values measured at 640 nm (Figure 3). Finally, we calculated the total amount of PACs based on a calibration curve prepared with a PAC-A standard using mixtures with increasing percentages of PAC-B and decreasing percentages of PAC-A at a final concentration of 20 µg/ mL, as calculated using the gravimetric method. Table 1 shows that the total amount of PACs increased with increasing PAC-B content despite a constant amount of total PACs used. These results indicate that the BL-DMAC method is time-sensitive (in our analyses, the best timing for the PAC-A reaction was 20 min) and that shorter reaction times may lead to overestimating the total PAC content. The latter result may occur in cases of high amounts of PAC-B. Furthermore, our results confirm that the BLDMAC method does not distinguish between PAC-A tion-to-treat principle was followed. RESULTS One of the key points in UTI prevention is the assessment of the content of bioactive PACs-A, which are contained in the capsule. Total PAC content can be quantitated using various methods, among which the BL-DMAC method is generally recognised as the most accurate.(12,13) However, the BL-DMAC assay is sensitive to the presence of both PACs-A and PACs-B. When we measured the standards of PAC-A and PAC-B using the BL-DMAC method, we found that the same amount of PAC reacted differently with various absorbance spectra (Figure 1). In particular, a higher absorbance (at 640 nm) was found for PAC-B than PAC-A (Figure 1). A time-course experiment was then performed by measuring the BL-DMAC reaction of 20 µg/mL PAC-A and PAC-B. PAC-B reached maximum absorbance after 11 min, whereas PAC-A showed a maximum absorbance at 20 min (Figure 2). PAC-A and PAC-B purity standards were analysed to assess purity and integrity; the results Miscellaneous 2645 Figure 5. Boxplot representing the logarithm of the scores of the placebo and Oximacro® groups in both females and males volunteers. Females differences between placebo and Oximacro®: Mann-Whitney = 875; P < .001; n = 60; males differences between placebo and Oximacro®: Mann-Whitney = 24; P = .016; n = 10. Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al. Figure 6. Boxplot representing the logarithm of the scores of the placebo and Oximacro® groups in both females (left panel) and males (right panel) volunteers according to the age range. Left panel, females age range-based (19-24, 25-30, 36-40, 41-50 and > 51 years) differences between placebo and
Oximacro®: for each age range, Mann-Whitney = 25; P = .008; n = 10; differences between placebo and Oximacro® for the female age range of 31-3
years: Mann-Whitney = 20.5; P = .095; n = 10. Right panel, differences between placebo and Oximacro® for the male age range over 51: Mann-Whitney
= 24; P = .016; n = 10.
and PAC-B and that even when a calibration curve is
obtained with PAC-A, an increased amount of PAC-B
eventually increases the absorbance at 640 nm, thereby
affecting the total PAC quantification. Therefore, a high
PAC value obtained with the BL-DMAC method does
not necessarily indicate a high amount of PAC-A even
though a calibration curve is calculated with a PAC-A
standard.
Having assessed the best timing for PAC determination, we next measured the total amount of Oximacro® PACs. The BL-DMAC method showed a total of
372.34 mg/g (± 2.3) PACs, in line with that declared
by the producer. To our knowledge, and based on the
BL-DMAC method, this is the highest amount of PACs
reported for a cranberry extract that is currently on the
market.
The bioactivity of cranberry against UTI is dependent
on the PAC-A content(9,15,16) (particularly dimers and
trimers);(17-19) thus, we analysed Oximacro® via HPLC
followed by electrospray ionization (EI) and tandem
mass spectrometry (ESI-MS/MS). Oximacro® was
primarily composed of PAC-A dimers followed by a
lower amount of PAC-A trimers (Figures 4a and 4c).
The total percentages of PAC-A and PAC-B based on
HPLC-ESI-MS/MS were 86.72% (± 1.65) and 13.99%
(± 1.03), respectively (Figures 4a-c). The percentage
of other PAC polymers was below the threshold of detection.
Considering the total PACs (calculated using the BLDMAC method) and the percentage of PAC-A dimers
and trimers, Oximacro® showed a total PAC-A content
of 322.89 mg/g (± 1.58). Therefore, 112 mg of Oximacro® contains 360 mg/g PAC-A. Following the assessment and authentication of the PAC-A content of
Oximacro®, we prepared capsules containing 112 mg
of Oximacro® (which corresponds to 36% PAC-A) and
placebo capsules. In all volunteers, the infection prior
to recruitment was due to E. coli in 85%, other enteric
Gram-negative bacilli in 10%, and more than 1 type of
bacteria in 5% of the volunteers. Table 2 shows the volunteers’ demographic and baseline characteristics.
Reasons for dropout in the experimental group included
relocation (1), feeling better prior to the end of treatment (4), contrary advice from a family doctor (1),
and a family perception of Oximacro® ineffectiveness
(1). Reasons for dropout in the placebo group included
acute pain (12), contrary advice from a family doctor
(2), and a family perception of Oximacro® ineffectiveness (1). The median follow-up time in both groups was
4 weeks.
The mean capsule intake was 97% (95% CI: 96.6–
97.6%) and was similar between the experimental and
placebo groups. After 7 days of Oximacro® and placebo administration, a contingency table was calculated
based on recovered vs. not recovered volunteers (Table
3); this table showed a significant difference (Fisher’s
exact test: P < .001) between the Oximacro® and placebo groups. A general Mann-Whitney U-test test further showed a highly significant difference between the placebo and Oximacro® groups (1191; P < .001; n = 70), with most of the placebo group unable to recover from UTI. Eventually, all placebo volunteers had to be treated with antibiotics (Monuril®, trometamol salt of fosfomycin) to reduce pain. Figure 5 shows the boxplot of female and male scores; a significant difference was found between the placebo and Oximacro® groups for both females (Mann-Whitney U-test = 875; P < .001; n = 60) and males (Mann-Whitney U-test = 24; P = .016; n = 10). Significant differences were also found when the age ranges were analysed. In particular, between the placebo and Oximacro® groups, the female age ranges of 19-24, 25-30, 36-40, 41-50 and over 51 years showed Vol 13 No 02 March-April 2016 2646 Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al. highly significant differences (for each age range: Mann-Whitney U-test = 25; P = .008; n = 10); the age range from 31-35 years showed barely significant differences (Mann-Whitney U-test = 20.5; P = .095; n = 10) (Figure 6 left panel). For males, the only age range (over 51) showed a significant difference between the placebo and Oximacro® groups as reported above (Figure 6 right panel). Finally, considering the CFU/mL counts from the urocultures, a significant difference (P < .001) was found in the comparison between the experimental group and the placebo group (SD difference = 51688; df = 34, t = -10.27; Dunn-Sidak Adjusted P < .001, Bonferroni Adjusted P < .001). Overall, these results show that the administration of Oximacro® significantly ameliorated UTI in the treatment group. When a multivariate binary logistic regression was performed to examine the independent effect of Oximacro® on healing (dependent variable) based on sex and age (covariates), no significant effect (P > .900)
for categorical variables was found for the treatment
outcomes (data not shown).
DISCUSSION
cranberry extract products via both BL-DMAC and
HPLC found that BL-DMAC values for the PAC content per unit were below those declared by the manufacturers. In particular, some cranberry extract medicinal
products showed a A-type PAC content so low that they
would have no chance of providing health benefits;(32)
the availability of these extracts was likely the result
of overestimation of the PAC content provided by the
Bates-Smith and the European Pharmacopoeia methods. On the one hand, these methods grant a high percentage value; on the other hand, by overestimating the
real PAC content, these methods limit the health benefits of cranberry extracts. The cranberry industry is currently using BL-DMAC as a standard method;(13) however, the BL-DMAC method is unable to discriminate
between A- and B-type PACs.(13) It is thus important to
combine accurate timing and the kinetics of the method
with the HPLC-MS authentication of PAC types. The
results of this work show that the presence of PAC-B in
cranberry extract can overestimate the total PAC content based on a PAC-A calibration curve. For instance,
a cranberry extract with a high percentage of PAC-B (as
is typical in some cranberry cultivars) may yield a high
total PAC value with the BL-DMAC method despite a
PAC-A standard calibration curve. Furthermore, in our
conditions (see the Materials and Methods), 20 min of
reaction were required for an accurate PAC-A determination (complete saturation of the reaction). Therefore, the standardization of PACs using the BL-DMAC
method and the authentication of PAC-A with LC-MS
is a prerequisite in preparing cranberry dosages for the
prevention of UTIs.
Our results showed that the cranberry extract Oximacro® contains a high total PAC content and a high percentage of bioactive PAC-A dimers and trimers. When
administered to volunteers, the extract was particularly
suitable for UTI prevention, and 112 mg Oximacro®
(equivalent of 36 mg PACs-A) twice per day for 7 days
was significantly effective in reducing the total urobacterial CFU counts in both the female and male groups
with respect to placebo. The age ranges were unaffected
by treatment with the sole exception of the 31-35 year
age range in the female group. This group did not differ
in baseline characteristics with respect to the other age
groups; thus, the reason for the reduced effect of Oximacro® in this group requires further investigation. A
literature search on age-related responses to cranberry
treatment did not provide any reported cases, although
further studies will focus on this aspect.
The efficacy of cranberry in preventing UTIs remains
controversial primarily due to contrasting results indicating either a nonsignificant effect (as in the case
of cranberry juice drinking)(20-23) or an extended duration therapy requirement (e.g., 12 months of drinking
cranberry juice).(24) In individuals with recurrent UTIs,
low-dose antibiotic prophylaxis for several months is
usually recommended.(25) However, extended use of
antibiotics may lead to the development of antibiotic
resistance. Indeed, several E. coli isolates are resistant
to antimicrobial treatment, and the interest in non-antibiotic methods for the prevention of UTIs is growing.(26)
If the dosage of non-antibiotic methods is not standardized, the cost/effect ratio may be higher than antibiotic
treatment, as recently shown with a cranberry prophylaxis regimen for preventing UTIs in which the PAC-A
treatment was far below (18.2 mg/day) the recommended dosage (72 mg). In this case, the cranberry treatment
was less effective and more expensive than (dominated
by) trimethoprim-sulfamethoxazole prophylaxis.(27) The
use of concentrated cranberry extract with a high PAC
content has been successfully proven to prevent UTIs
in women who are subject to recurrent infections.(7,9,10,
18,28-31)
Despite these positive results, one of the major
concerns is the quantification of PACs-A, which are the
only bioactive compounds thus far demonstrated to exert a significant uropathogenic bacterial anti-adhesion
effect.(9,19) A recent survey on the PAC content of some CONCLUSIONS
Miscellaneous 2647
Prevention of Urinary Tract Infection with Oximacro®-Occhipinti et al.
The results of this work are in agreement with previous
randomized, double-blind versus placebo multicentre
studies examining the effects of 72 mg of PAC-standardized cranberry.(2,10,13) Furthermore, our results show
that 72 mg PAC-A is highly effective, and we suggest
the use of dosages based on PACs-A instead of the total
PACs in UTI treatment. Due to the impossibility of BLDMAC in discriminating between PAC-A and PAC-B,
the sole total PACs quantification may not be sufficient
in providing the required amount of PAC-A needed to
significantly inhibit UTIs. Further studies will assess
the recurrence of UTIs in Oximacro®-supplemented
volunteers.
AKNOWDLEDGEMENTS
The authors would like to express their gratitude to Dr.
Francesca Barbero for the statistical analyses and for
the revision of the manuscript.
CONFLICTS OF INTEREST
None declared
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Miscellaneous 2649
Consumption of a cranberry juice beverage lowered the number of
clinical urinary tract infection episodes in women with a recent history
of urinary tract infection1
Kevin C Maki,2,3* Kerrie L Kaspar,4 Christina Khoo,4 Linda H Derrig,2 Arianne L Schild,2 and Kalpana Gupta5,6
ABSTRACT
Background: Urinary tract infections (UTIs) are among the most
common bacterial infections and are often treated with antibiotics.
Concerns about multidrug-resistant uropathogens have pointed to
the need for safe and effective UTI-prevention strategies such as
cranberry consumption.
Objective: We assessed the effects of the consumption of a cranberry beverage on episodes of clinical UTIs.
Design: In this randomized, double-blind, placebo-controlled, multicenter clinical trial, women with a history of a recent UTI were
assigned to consume one 240-mL serving of cranberry beverage/
d (n = 185) or a placebo (n = 188) beverage for 24 wk. The primary
outcome was the clinical UTI incidence density, which was defined
as the total number of clinical UTI events (including multiple events
per subject when applicable) per unit of observation time.
Results: The dates of the random assignment of the first subject and
the last subject’s final visit were February 2013 and March 2015,
respectively. The mean age was 40.9 y, and characteristics were
similar in both groups. Compliance with study product consumption
was 98%, and 86% of subjects completed the treatment period in
both groups. There were 39 investigator-diagnosed episodes of clinical UTI in the cranberry group compared with 67 episodes in the
placebo group (antibiotic use–adjusted incidence rate ratio: 0.61; 95%
CI: 0.41, 0.91; P = 0.016). Clinical UTI with pyuria was also significantly reduced (incidence rate ratio: 0.63; 95% CI: 0.40, 0.97; P =
0.037). One clinical UTI event was prevented for every 3.2 womanyears (95% CI: 2.0, 13.1 woman-years) of the cranberry intervention.
The time to UTI with culture positivity did not differ significantly
between groups (HR: 0.97; 95% CI: 0.56, 1.67; P = 0.914).
Conclusion: The consumption of a cranberry juice beverage lowered
the number of clinical UTI episodes in women with a recent
history of UTI. This study was registered at clinicaltrials.gov
as NCT01776021.
Am J Clin Nutr 2016;103:1434–42.
Keywords: antibiotics, bacteria, cranberry, inflammation,
proanthocyanidin, urinary tract infection, women
INTRODUCTION
A urinary tract infection (UTI) is common and increasingly
difficult to treat because of the rising rates of antibiotic resistance (1, 2). Approximately 60% of women will experience $1
1434
UTI in their lifetimes, and UTIs are responsible for w10.5
million physician office and emergency department visits annually in the United States (3, 4). The costs attributable to UTIs
include those for antibiotic therapy, visits to health care providers, laboratory testing, and lost productivity (1). It has been
estimated that 25–35% of women diagnosed with a UTI will
suffer a recurrence within 6 mo (1, 5). The prevention of a UTI
is most effectively achieved with antibiotic prophylaxis (6), although it has been recommended that, in women with recurrent
cystitis, prophylactic antimicrobial therapy should be used only
when nonantimicrobial therapy is not effective (7, 8). Increasing
rates of antibiotic resistance (9–12) and other adverse effects from
antibiotic exposure make this approach important to consider.
Cranberry consumption has been evaluated as a strategy for
reducing clinical UTI recurrence in women with a recent history
of a UTI (5, 13). Results from randomized clinical trials have
been generally suggestive of a benefit but have often lacked
sufficient statistical power to provide definitive results (14). The
current study was conducted to compare the effects of the
consumption of a cranberry beverage with that of a placebo
beverage on the clinical (symptomatic) UTI incidence density in
healthy women with a recent history of a UTI.
METHODS
Study design
The study was a 24-wk multicenter, double-blind, randomized,
placebo-controlled trial that was designed to assess the effects of
the consumption of a cranberry beverage on episodes of clinical
(symptomatic) UTI in healthy women. The study was conducted
at 17 clinical research sites in the United States and at one clinical
1
Supported by Ocean Spray Cranberries Inc. Beverages were provided by
Ocean Spray Cranberries Inc. This is a free access article, distributed under
terms (http://www.nutrition.org/publications/guidelines-and-policies/license/)
that permit unrestricted noncommercial use, distribution, and reproduction
in any medium, provided the original work is properly cited.
*To whom correspondence should be addressed. E-mail: kmaki@
mbclinicalresearch.com.
Received January 11, 2016. Accepted for publication March 17, 2016.
doi: 10.3945/ajcn.116.130542.
Am J Clin Nutr 2016;103:1434–42. Printed in USA. Ó 2016 American Society for Nutrition
Downloaded from https://academic.oup.com/ajcn/article/103/6/1434/4569625 by guest on 24 April 2022
2
Biofortis Clinical Research, Addison, IL; 3MB Clinical Research, Glen Ellyn, IL; 4Ocean Spray Cranberries, Lakeville-Middleboro, MA; 5Department of
Medicine, Boston University School of Medicine, Boston, MA; and 6VA Boston Healthcare System, Boston, MA
1435
CRANBERRY JUICE AND CLINICAL UTI
research site in France between February 2013 and March 2015.
The protocol was approved by an institutional review board in the
United States (Quorum Review IRB, Seattle, Washington) and by
the National Security Agency for Medicines and Health Products
and an Ethical Research Committee (Committee for Personal
Protection) in France. Procedures were followed in accordance
with the Declaration of Helsinki of 1975 as revised in 1983.
Written informed consent was obtained from all subjects.
Subjects
Study products
Subjects were randomly assigned (1:1 ratio) to consume one
8-oz (240-mL) bottle of cranberry or placebo study beverage per
day throughout the 24-wk treatment period. The randomization
sequence was generated with SAS for Windows software (version
9.1.3; SAS Institute Inc.) by a blinded statistician with the use of
a seed number and random allocation in blocks by research site.
The randomization module of the DATATRAK Electronic Data
Capture system (DATATRAK ONE UX, versions 13.0.0 to
13.3.5) was used for coded treatment allocation at each research
site by a study coordinator or investigator.
Beverages were provided by Ocean Spray Cranberries Inc. and
were stored at room temperature and refrigerated before consumption. Cranberry and placebo beverages each provided
w35 kcal (w146 kJ)/240-mL serving. The placebo beverage
contained filtered water, fructose, dextrose, citric acid, quinic acid,
malic acid, natural flavors, pectin, potassium citrate, sodium citrate,
red 40, blue 1, acesulfame-potassium, and sucralose. The active
study beverage contained filtered water, cranberry juice from concentrate, fructose, natural flavors, pectin, sodium citrate, acesulfamepotassium, and sucralose. The analytic composition of the study
TABLE 1
Study-product composition1
Component
2
Energy, kcal
Carbohydrates,3 g
Sugars,4 g
Organic acids,6 g
Vitamin C,7 mg
Proanthocyanidins, mg
DMAC-method I8
DMAC-method II9
Anthocyanins,4 mg
Phenolic acids,10 mg
Flavanols and flavonols,10 mg
Total phenolics,11 mg
1
2
Cranberry beverage
Placebo beverage
36
9
6.3 6 0.15
2.0 6 0.1
ND
34
8
6.9 6 0.3
1.9 6 0.2
ND
6
6
6
6
6
6
ND
ND
ND
ND
ND
17.0 6 5.4
41.1
119
1.3
5.4
8.3
135
7.1
16.9
0.8
0.8
3.6
30.7
DMAC, 4-dimethylaminocinnamaldehyde; ND, not detected.
Determined with the use of Atwater factors (Covance Laboratory)
(19).
3
Determined with the use of the carbohydrate by difference method
(20).
4
Measured with the use of HPLC (21).
Mean 6 SD where applicable per 240-mL serving (all such values).
6
Measured with the use of ion chromatography (21).
7
Measured with the use of an iodiometric titration method (22).
8
Determined with the use of a DMAC colorimetric method with
procyanidin A2 as a standard (23).
9
Determined with the use of a modified DMAC colorimetric method
with the cranberry proanthocyanidin standard isolated and purified from
cranberry juice concentrate (21, 24).
10
Determined with the use of HPLC (25).
11
Determined with the use of the Folin-Ciocalteu reagent colorimetric
method (21).
5
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Eligible subjects included women who were 20–70 y of age
with BMI (in kg/m2) ,40.0 and a recent history of a UTI, which
was defined as $2 episodes of a UTI that were treated by
a health care professional in the past year (self-report) of which
$1 UTI had been treated #6 mo of the screening visit. Women
who were using prophylactic antibiotics for a UTI were not
enrolled, but a 2-wk washout period from antibiotic use was
allowed before screening. Individuals with an active infection or
signs or symptoms of a UTI or other active infection were excluded. If a clean-catch urine sample at screening was positive
for asymptomatic bacteriuria ($105 CFU for a uropathogen), the
woman was rescreened $2 wk later.
Subjects agreed to avoid the consumption of Vaccinium
products (blueberries, cranberry juice, cranberries, dried cranberries, and cranberry or blueberry powders, pills, or supplements) and probiotic dietary supplements and to limit the
consumption of all probiotic-containing foods or yogurt, soda,
and energy drinks within 2 wk before screening and through
week 24. The daily consumption of carbonated beverages and
fermented milk products has been associated with a reduced risk
of recurrent UTI in some studies, although this finding has not
been universal (15–18). In the current study, subjects were allowed to consume carbonated beverages, energy drinks, and
yogurt but were asked to avoid intakes that were far above mean
US intakes to allow the results to be generalizable while minimizing the potential for confounding by extreme intakes of these
products. Subjects received a stipend for their participation in
the study, which included 5 clinic visits (screening and baseline
visits at weeks 21 and 0, respectively, and 3 treatment visits at
weeks 8, 16, and 24) and 9 telephone contacts at weeks 2, 4, 6,
10, 12, 14, 18, 20, and 22 to improve study compliance by reminding subjects to record data in their daily diaries.
Individuals were excluded from participation in the study if
they used a bladder catheter or had polycystic disease, interstitial
cystitis, previous urologic surgery, stones, anatomical abnormalities of the urinary tract, a spinal cord injury, conditions that
produce immunocompromise, severe renal impairment, or
multiple sclerosis. Additional exclusionary conditions included
diabetes mellitus with glycated hemoglobin $8.0%, diabetes
mellitus treated with insulin, a history or presence of cancer in
the previous 2 y (except nonmelanoma skin cancer), a recent
(within the past 3 mo) major trauma or surgical event, or the use
of oral anticoagulants #4 wk before screening. Women were
also excluded from participation if they had an abnormal laboratory test of clinical importance. Women who were pregnant,
planning to be pregnant during the study, or lactating were excluded from the study, and women of childbearing potential had
to commit to the use of a medically approved form of contraception throughout the study.
Subjects were instructed to maintain a stable body weight, adhere to habitual exercise patterns, and avoid the consumption of
foods that are high in polyphenols for the 24 h before and during the
24-h urine collection periods. Cigarette smokers were instructed to
abstain from tobacco products 1 h before and during clinic visits.
1436
MAKI ET AL.
Measurements
Subjects completed a validated daily diary in which they recorded
their consumption of the study beverage and captured any UTI
symptoms (26). Daily diaries were reviewed at each post–random
assignment clinic visit, and subjects were queried during telephone
calls between clinic visits regarding their compliance with the
consumption of the study product as well as whether symptoms or
adverse experiences had occurred. If symptoms occurred at any
time during the study, the subject was instructed to call the research
clinic to arrange for a UTI-evaluation visit, which included a pelvic
examination. A clean-catch urine sample was also collected, and
a clinical (symptomatic) UTI was diagnosed by the investigator on
the basis of $1 of the following symptoms: dysuria, urinary frequency, urinary urgency, or suprapubic pain in the absence of other
potential etiologies such as vaginal infection or discharge. Investigators treated a clinical UTI with a standardized antimicrobial
therapy regimen, and subjects continued to consume the study
beverages during treatment. Clean-catch urine samples were also
collected at weeks 21, 0, 8, 16, and 24 for urinalysis and culture.
The presence or absence of pyuria in clean-catch urine samples
was determined by a leukocyte esterase dipstick result (27). In the
United States, the urine culture at the screening visit and the urinalysis from all visits were analyzed by Johnson City Medical
Center (Johnson City, Tennessee), and urine cultures from nonscreening visits were tested by The General Clinical Research Center (University of Washington, Seattle, Washington).
In France, the urine culture at the screening visit and urinalysis
from all visits were analyzed by Synevo Central Laboratory
Poland (Gdansk, Poland), and urine cultures from nonscreening
visits were tested in Barcelona, Spain, by Servei de Microbiologia, Hospital Universitari Vall d’Hebron. At weeks 0, 8,
16, and 24, subjects completed questionnaires that assessed
sexual history, food and beverage consumption, and the presence
and severity of gastrointestinal symptoms.
Statistical analyses
The primary outcome variable was the clinical (symptomatic)
UTI incidence density, which was defined as the number of
clinical UTI events in each group (including multiple events per
subject when applicable) per unit of observation time. The incidence density was selected as the primary outcome variable
because UTI episodes often cluster in time (28, 29). The selection
of a clinical UTI as the primary outcome in the current study was
consistent with guidelines for UTI management (30) because
initial treatment decisions are generally made before the availability of culture results, and the presence of multiple symptoms
in the absence of vaginal discharge in women with a history of
UTI is highly predictive (UTI probability .90%) (7, 31).
The following 3 classifications were used for clinical UTI
analyses: investigator-diagnosed UTIs, probable UTIs, and
possible UTIs. Investigator-diagnosed clinical UTIs were those
for which the investigator evaluated the subject and made the UTI
diagnosis. Probable UTIs were those for which the investigator
did not examine the subject, but a nonstudy health care provider
did examine the subject and prescribed antibiotics. Possible UTIs
were those that did not fall into either of the other 2 categories
including those in which the subject self-treated and instances
when it was not clear whether an episode was a new UTI or
a continuation of a previous infection that had not cleared (e.g.,
when a subject’s symptoms stopped and recurred within 2 wk
with no intervening test-of-cure visit). Results for investigatordiagnosed UTIs are presented in detail, and results from analyses that included probable and possible UTIs are described as
sensitivity analyses.
Secondary and exploratory outcome variables included the
incidence density for a clinical UTI with pyuria, the time from
random assignment to a first clinical UTI, the time from random assignment to a first clinical UTI with pyuria, and the time
from random assignment to a first symptomatic UTI with culture positivity ($10 3 CFU/mL) for any uropathogen and
for Escherichia coli. Safety was assessed by an evaluation of
treatment-emergent adverse events, the frequency and severity
of gastrointestinal signs and symptoms, and the measurement of
vital signs, body weight, and clinical laboratory values.
Fisher’s exact test with the use of G*Power software (free
software available at http://www.gpower.hhu.de) was used to
complete power calculations (32). Because the incidence density
was to be used as the primary outcome variable, it was anticipated that the study power would be greater than that reflected
by this calculation (29), which did not account for the occurrence of multiple UTIs during the treatment period in some
women. On the basis of the assumption that the proportion of
women who would experience a UTI episode in the placebo
group would be 32% (5), a sample size of 145 subjects/group
was expected to provide $80% power (a = 0.05; 2 sided) to
detect a reduction to 17.8% in the treatment group. An enrollment of 300 subjects (150 subjects/group) was initially planned,
but a decision was later made to increase the enrollment by an
additional 40 subjects because a blinded review showed that the
rate of a first UTI was slightly below initial projections.
Statistical analyses were conducted with the use of SAS for
Windows software (version 9.1.3). All tests for significance were
performed at a = 0.05 (2 sided). The baseline comparability
of treatment groups for subject characteristics was assessed
with the use of ANOVA and chi-square tests. Analyses were
performed in all randomly assigned subjects (intent-to-treat
population) with the observation time censored at the time
that the study product was discontinued for subjects who did
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beverages is shown in Table 1 (19–25). The placebo beverage was
designed to look, smell, and taste like the cranberry beverage (27%
juice), and a separate sensory study showed no difference in the
proportions of subjects who correctly guessed if they were randomly
assigned to receive either the cranberry beverage (51%) or the
placebo beverage (40%) (n = 167; parallel design; P = 0.20). The
cranberry juice cocktail study beverage used was similar to
commercially available low-calorie products in its juice content
(27% cranberry juice) although additional measures were taken
to minimize the variability in the contents of proanthocyanidins
and other bioactives. These additional measures included production from a single lot of cranberry concentrate and the use of
a shorter time to expiration than is used for beverages that are
produced for commercial use.
Compliance with study beverage consumption was assessed by
having subjects return all unused bottles of dispensed study
product and empty bottles of study product that had been consumed to the clinic. This information was checked against diary
data, and subjects were queried to evaluate any discrepancies.
CRANBERRY JUICE AND CLINICAL UTI
RESULTS
A total of 373 subjects were randomly assigned to consume the
cranberry beverage (n = 185) or placebo beverage (n = 188), and
322 subjects [cranberry: n = 160 (86.5%); placebo: n = 162
(86.2%)] completed through week 24 of the study (Figure 1). Two
subjects in the cranberry group were randomly assigned in error
(one subject had asymptomatic bacteriuria and another subject did
not have sufficient literacy to understand the consent form). Both
subjects were discontinued from treatment once the errors were
discovered. Three subjects withdrew consent because of adverse
events that were unrelated to the treatment (cranberry group: oral
thrush and dizziness; placebo group: stomach complaints).
Demographic and baseline characteristics are shown in Table
2. Subjects had a mean age of 40.9 y, and the majority of subjects were white (67.0%) and of non-Hispanic or non-Latino
ethnicity (75.6%). The mean 6 SEM compliance with daily
study-beverage consumption was 98.1% 6 0.6% and 98.2% 6
0.5% in the cranberry and placebo groups, respectively.
A total of 53 UTI-assessment visits were completed for
subjects in the cranberry group, and 82 UTI-assessment visits
were completed in the placebo group. These visits resulted in the
diagnosis by study investigators of 39 clinical UTIs in the
cranberry group and 67 clinical UTIs in the placebo group (Table 3).
The fractions of clinical UTI diagnoses for which the subject
reported $2 UTI symptoms at the assessment visits were 97.4%
(38 of 39 diagnoses) for the cranberry condition and 99.0% (66 of
67 diagnoses) for the placebo condition, and for $3 UTI symptoms at the assessment visits, these values were 97.4% (38 of 39
diagnoses) for the cranberry condition and 91.0% (61 of 67 diagnoses) for the placebo condition. The annualized UTI incidence
density was significantly reduced in the cranberry compared with
FIGURE 1 Subject disposition throughout the trial (Consolidated Standards of Reporting Trials flow diagram).
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not complete the full treatment period. Assumptions of normality of residuals were investigated for each response measurement. In cases in which the normality assumption was
rejected at the 1% level with the use of the Shapiro-Wilk test
(33), an analysis with the use of ranks was performed.
The UTI incidence density was analyzed with the use of
Poisson regression with terms of treatment, site, country (United
States or France), time since last UTI category (#30, 31–89, or
$90 d), and age category (,50 or $50 y) as well as an offset
variable for the log of time (woman-years) under observation.
The model was reduced until treatment and any significant
terms (P , 0.05) remained with the use of a backward-elimination
method. The appropriate fit and overdispersion were assessed
for each model (34), and comparisons of results from full and
reduced models were completed to assess whether the modelreduction procedure materially altered the point estimates and
95% CIs for the treatment effect. An adjustment for the susceptible time under observation to account for antibiotic use was
calculated with the use of a subtraction of 7 d from the susceptible time for each instance of antibiotic use regardless of the
reason for use. The incidence density is presented with and
without this adjustment.
Time-to-event outcome variables were analyzed with the use
of Cox proportional hazards models with the same covariates and
approach as previously described. HRs with 95% CIs and model
variable P values were determined. Assumptions of a constant
relative hazard were verified with the use of the Schoenfeld
residuals goodness-of-fit test (35). Continuous variables were
analyzed with the use of an ANCOVA and a model with a term
for the treatment and baseline included as a covariate.
1437
1438
MAKI ET AL.
TABLE 2
Demographic and baseline characteristics of subjects receiving cranberry or placebo beverages1
Characteristic
Placebo group (n = 188)
40.9 6 1.1
41.0 6 1.0
131 (70.8)
54 (29.2)
133 (70.7)
55 (29.3)
122
30
0
3
2
3
25
(65.9)
(16.2)
(0.0)
(1.6)
(1.1)
(1.6)
(13.5)
128
29
2
7
1
0
21
(68.1)
(15.4)
(1.1)
(3.7)
(0.5)
(0.0)
(11.2)
25
136
24
1.6
(13.5)
(73.5)
(13.0)
6 0.1
21
146
21
1.7
(11.2)
(77.7)
(11.2)
6 0.1
17
80
88
4.9
(9.2)
(43.2)
(47.6)
6 0.4
21
79
88
5.7
(11.2)
(42.0)
(46.8)
6 0.6
58
123
2
2
27.0
(31.4)
(66.5)
(1.1)
(1.1)
6 0.4
50
135
3
6
26.5
(26.6)
(71.8)
(1.6)
(3.2)
6 0.4
2
P
0.896
0.999
0.329
0.661
79
57
49
0.5
(42.7)
(30.8)
(26.5)
(0.0, 10.0)
134 (72.4)
19 (10.3)
32 (17.3)
92
40
56
0.4
(48.9)
(21.3)
(29.8)
(0.0, 12.0)
0.647
0.837
0.434
0.399
0.284
0.346
0.116
0.962
0.282
145 (77.1)
11 (5.9)
32 (17.0)
1
Baseline comparability of treatment groups for subject characteristics was assessed with the use of ANOVA and chisquare tests. UTI, urinary tract infection.
2
Mean 6 SEM (all such values).
3
Race and ethnicity were self-reported by subjects as part of a medical history questionnaire that was completed at
screening to allow comparison with the racial and ethnic compositions of the country.
4
For clinical studies in France, data regarding ethnicity can only be collected if justified by the type of research. In this
study of cystitis, there was no justification to investigate ethnicity, and therefore, this information was not collected from
subjects enrolled at the clinical site in France.
5
Previous history of a medical diagnosis of diabetes as recorded in the medical history. In the cranberry group, 2
subjects with diabetes were $50 y of age; in the placebo group, 2 subjects with diabetes were ,50 y of age, and 4 subjects
were $50 y of age.
6
All values are medians; minimums, maximums in parentheses. These data were not normally distributed (normality
assumption was rejected at the 1% level with the use of the Shapiro-Wilk test) and were ranked in the analyses.
in the placebo arm (incidence rate ratio: 0.62; 95% CI: 0.42, 0.92;
P = 0.017). Adjustment for antibiotic use (139 instances in the
control group and 111 instances in the cranberry group) for the
susceptible time under observation did not materially alter the
point estimate (incidence rate ratio: 0.61; 95% CI: 0.41, 0.91; P =
0.016). For every 3.6 woman-years (95% CI: 2.3, 15.8 womanyears) of the cranberry intervention, 1 symptomatic UTI event
was prevented. After adjustment for antibiotic use, 1 symptomatic
UTI was prevented for every 3.2 woman-years (95% CI: 2.0, 13.1
woman-years).
Results were not materially influenced by the model reduction
(data not shown). There was no statistical heterogeneity in the
treatment response for subjects ,50 and $50 y of age (P-treatment
by age-group interaction = 0.526).
The incidence density for symptomatic UTIs with pyuria
(adjusted for antibiotic use) was significantly reduced in the
cranberry arm compared with in the placebo arm (Table 3)
(incidence rate ratio: 0.63; 95% CI: 0.40, 0.97; P = 0.037). With
the use of the classifications of investigator-diagnosed plus
probable symptomatic UTIs (47 and 72 UTIs in the cranberry
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Age, y
Age subgroup, y, n (%)
,50
$50
Race,3 n (%)
White
Black/African American
American Indian/Alaskan Native
Asian or Pacific Islander
Multiracial origin
Other
Missing4
Ethnicity, n (%)
Hispanic/Latino
Not Hispanic/Latino
Missing
Treated UTIs in past 6 mo, n
Most recent UTI history, d, n (%)
#30
31–89
$90
Vaginal intercourse frequency in past 4 wk, n
Sexual partners in past 4 wk, n (%)
0
1
$2
History of diabetes,5 n (%)
BMI, kg/m2
Categorical BMI, kg/m2, n (%)
,25
25 to ,30
$30
Alcoholic drinks,6 n/wk
Smoking status, n (%)
Nonsmoker
Current smoker
Past smoker
Cranberry group (n = 185)
1439
CRANBERRY JUICE AND CLINICAL UTI
TABLE 3
Symptomatic episodes of UTI diagnosed an…